The induction of the hepatic, microsomal, mixed function oxidases by phenobarbital has been extensively studied in the whole animal. But studies of this phenomenon have been limited in the isolated hepatocyte because of a loss of the activity during culture. In the past year we have been able to demonstrate the maintenance of this activity requires the presence of insulin, thyroxine, and dexamethasone as well as 20% serum from adult, male rats. An initial objective of this study shall be to optimize the culture condition in an effort to maintain mixed function oxidase activity in the absence of serum. We propose to investigate optimization of hormones, addition of gastrointestinal hormones such as secretin, cholecystokinin, glucagon, and gastrin in maintaining activity. Secondly, we propose to examine the possible effects of various factors as Se and vitamin E in maintaining activity through maintenance of the cellular mechanism, for protection against oxygen free radicals. During the course of our studies on the isolated hepatocyte we have also used them for short term studies. We have observed an effect of oxygen on the kinetic parameters of ethylmorphine metabolism which imply that in the projection of in vitro kinetic data to in vivo studies, the lower oxygen pressure of the liver must be taken into account. Also in these acute studies we have observed that the hepatocyte appears to actively transport ethylmorphine. We propose to extend the acute studies to an agent of greater therapeutic interest, phenytoin.